In JoVE (1)
Other Publications (8)
- The Biochemical Journal
- The Journal of Biological Chemistry
- International Journal of Cancer. Journal International Du Cancer
- Developmental Cell
- Clinical and Experimental Pharmacology & Physiology
- Naunyn-Schmiedeberg's Archives of Pharmacology
- Plant Physiology and Biochemistry : PPB / SociÃ©tÃ© FranÃ§aise De Physiologie VÃ©gÃ©tale
- Nature Medicine
Articles by I-Chun Tsai in JoVE
在体内病态的人类基因组建模使用斑马鱼 Adrienne R. Niederriter1,2, Erica E. Davis1,3, Christelle Golzio1, Edwin C. Oh1, I-Chun Tsai1, Nicholas Katsanis1 1Center for Human Disease Modeling, Department of Cell Biology, Duke University Medical Center, 2Department of Evolutionary Anthropology, Duke University, 3Department of Pediatrics, Duke University Medical Center 在这里，我们提出了一个系统的方法来开发有关生理，敏感，特异
Other articles by I-Chun Tsai on PubMed
Degradation of Human Thymidine Kinase is Dependent on Serine-13 Phosphorylation: Involvement of the SCF-mediated Pathway The Biochemical Journal. Feb, 2003 | Pubmed ID: 12435275 The expression level of human thymidine kinase (hTK) is regulated in a cell-cycle-dependent manner. One of the mechanisms responsible for the fluctuation of TK expression in the cell cycle can be attributed to protein degradation during mitosis. Given the facts that cell-cycle-dependent proteolysis is highly conserved in all eukaryotes and yeast cells are an excellent model system for protein-degradation study, here we report on the use of Saccharomyces cerevisiae and Schizosaccharomyces pombe to investigate the degradation signal and mechanism required for hTK degradation. We found that the stability of hTK is significantly reduced in mitotic yeasts. Previously, we have observed that Ser-13 is the site of mitotic phosphorylation of hTK in HeLa cells [Chang, Huang and Chi (1998) J. Biol. Chem. 273, 12095-12100]. Here, we further provide evidence that the replacement of Ser-13 by Ala (S13A) renders hTK stable in S. pombe and S. cerevisiae. Most interestingly, we demonstrated that degradation of hTK is impaired in S. cerevisiae carrying a temperature-sensitive mutation in the proteasomal gene pre1-1 or the Skp1-Cullin-1/CDC53-F-box (SCF) complex gene cdc34 or cdc53, suggesting the contribution of the SCF-mediated pathway in hTK degradation. As phosphorylation is a prerequisite signal for SCF recognition, our results implied that phosphorylation of Ser-13 probably contributes to the degradation signal for hTK via the SCF-mediated proteolytic pathway.
酪蛋白激酶的调节我 ε 活动由 Wnt 信号。 The Journal of Biological Chemistry. Mar, 2004 | Pubmed ID: 14722104 连环 Wnt/β-蛋白信号转导通路是重要的发展和癌症。酪蛋白激酶 Iepsilon (CKIepsilon) 是典型的 wnt 信号转导通路的积极监管机构。CKIepsilon 本身可以受体外抑制自磷酸化，和最近的数据表明在体内的激酶活性可以受胞外刺激。我们在这里展示磷酸化状态和激酶活性的 CKIepsilon 直接受规管的 Wnt 信号。共表达的 XWnt-8 或另外的可溶性 Wnt-3a 配体导致活性的内源性 CKIepsilon 重大和迅速增加。CKIepsilon 活动的增加是下降抑制自磷酸化的结果，因为它由 preincubation immunoprecipitated 激酶与 ATP 的废除。此外，突变的 CKIepsilon 抑制自磷酸化站点创建称为 CKIepsilon(MM2)，是明显比 CKIepsilon 更活跃，不会进一步激活激酶后 Wnt 刺激。Autoinhibition CKIepsilon 是生物相关的因为 CKIepsilon(MM2) 是比在激活从 Lef1 依赖的启动子转录 CKIepsilon 更有效。最后，在非洲爪蟾胚胎中的 CKIepsilon(MM2) 表达诱导轴重复和额外的发育异常。数据表明 Wnt 信号通过造成瞬态除磷的关键抑制站点中的激酶的羧基末端域存在激活 CKIepsilon。经典 wnt 信号通路的激活可能因此刺激细胞磷酸酶使其和激活 CKIepsilon
疾病关联的酪蛋白激酶我三角洲突变可能促进性腺瘤性息肉形成通过 Wnt 连环独立机制。 International Journal of Cancer. Journal International Du Cancer. Mar, 2007 | Pubmed ID: 17131344 Wnt 信号通路是胚胎发育的关键，是在多个癌症水平及。两个密切相关的异构体的酪蛋白激酶 （CKIdelta 和 ε） 是此途径的积极监管机构。我们推测突变 autoinhibitory 域中的 CKIdelta/ε 可能长期 CKIdelta/ε 活动和因此 Wnt 信号转导和增加腺瘤性息肉和患结肠癌的风险。外显子编码的 CKIepsilon 和 CKIdelta 的管理域被从从有腺瘤性息肉的个人和家族史的结肠癌患者的家族性腺瘤性息肉病不会影响或遗传性非息肉病性结直肠癌 （HNPCC) 获得的 DNA 测序。CKIdelta 错义突变，更改一个高保守的残留物，Arg324，为他 （R324H），发现在个人与大型及多发性息肉诊断年龄相对较小。两项调查结果表明这种突变是生物活性。第一、 异位腹侧表达式的 CKIdelta(R324H) 在非洲爪蟾胚胎在次坐标轴的形成与表附加特色型 (改变形态运动) 与不受管制的 CKIepsilon 见到的类似的结果。第二，CKIdelta(R324H) 是比 wildtype CKIdelta RKO 结肠癌细胞转型中的更有效。虽然 R324H 突变不会显著更改 CKIdelta 激酶活性体外激酶含量测定或 Wnt/β-连环蛋白信号转导所评估的一种 β-连环蛋白记者检测方法中，它会改变形态发生运动通过在非洲爪蟾早期发展中的 β-连环蛋白-独立机制。这种新的人类 CKIdelta 突变可能改变的生理作用和加强 CKIdelta 通过 Wnt 连环独立机制的转型能力，从而影响了结肠腺瘤发展。
Wnt-CKIvarepsilon-Rap1 通路调控 Gastrulation 通过调节 SIPA1L1，激活蛋白的 Rap 蛋白。 Developmental Cell. Mar, 2007 | Pubmed ID: 17336901 非经典 Wnt 信号在脊椎动物 gastrulation 期间控制形态发生运动。酪蛋白激酶我 ε (CKIvarepsilon) 是 Wnt 激酶调控 Wnt/β-连环蛋白信号并具有 β-连环蛋白-独立角色中了解得很少的形态发生。我们这里报告的 CKIvarepsilon 的绑定标识合作伙伴，SIPA1L1/E6TP1，说唱小蛋白家族的差距 (蛋白激活蛋白)。我们展示 CKIvarepsilon phosphorylates SIPA1L1，减少它的稳定，从而增加 Rap1 激活。Wnt-8，激活 CKIvarepsilon，增强了的 CKIvarepsilon 依赖的磷酸化和降解的 SIPA1L1。在早期非洲爪蟾或斑马鱼发展，灭活 Rap1 通路的结果异常 gastrulation 和缩短的前后轴。虽然 CKIvarepsilon 还传感器 Wnt/β-连环蛋白信号转导，Rap1 的抑制作用不会改变 β-连环蛋白调控基因的表达。我们的数据在规范化 Wnt 信号为 CKIvarepsilon 示范作用，并表明 Wnt 调控形态发生部分通过介导 CKIvarepsilon Rap1 信号控制。
Pipoxolan Inhibits Proliferation of HL-60 Human Leukaemia Cancer Cells by Arresting the Cell Cycle at the G0/G1 Phase Clinical and Experimental Pharmacology & Physiology. May, 2010 | Pubmed ID: 20082627 1. The aim of the present study was to investigate the molecular mechanisms by which pipoxolan exerts its inhibitory effects and apoptotic activity in human leukaemia HL-60 cells. 2. The effects of pipoxolan on the proliferation of HL-60 cells and on the distribution of cells within different phases of the cell cycle were investigated indirectly using a Trypan blue assay and a flow cytometer, respectively. The effects of pipoxolan on the apoptosis of HL-60 cells was investigated using DNA fragmentation and flow cytometer. The expression of factors affecting the cell cycle and apoptosis, including p53, p21, Bax, Bcl2, cytochrome c, caspase 3 and caspase 9, was examined by western blotting. 3. At 6.25 microg/mL, pipoxolan significantly induced apoptosis in human leukaemia HL-60 cells after 24 h exposure. In addition, HL-60 cells were arrested in the G(0)/G(1) phase via the induction of p53/p21 by pipoxolan. Apoptosis was associated with an increased Bax/Bcl-2 ratio, cytochrome c release, cleavage of procaspases-9 and -3 and hydrolysis of poly(ADP-ribose) polymerase. Intracellular reactive oxygen species (ROS) seem to play a key role in the pipoxolan-induced apoptosis, because high levels of ROS were produced early in the drug treatment. Apoptosis was significantly abrogated by the free radical scavenger N-acetylcysteine (NAC).
NP-184[2-(5-methyl-2-furyl) Benzimidazole], a Novel Orally Active Antithrombotic Agent with Dual Antiplatelet and Anticoagulant Activities Naunyn-Schmiedeberg's Archives of Pharmacology. Jun, 2010 | Pubmed ID: 20349046 The established antiplatelet and anticoagulant agents show beneficial effects in the treatment of thromboembolic diseases; however, these drugs still have considerable limitations. The effects of NP-184, a synthetic compound, on platelet functions, plasma coagulant activity, and mesenteric venule thrombosis in mice were investigated. NP-184 concentration-dependently inhibited the human platelet aggregation induced by collagen, arachidonic acid (AA), and U46619, a thromboxane (TX)A(2) mimic, with IC(50) values of 4.5 +/- 0.2, 3.9 +/- 0.1, and 9.3 +/- 0.5 microM, respectively. Moreover, NP-184 concentration-dependently suppressed TXA(2) formations caused by collagen and AA. In exploring effects of NP-184 on enzymes involved in TXA(2) synthesis, we found that NP-184 selectively inhibited TXA(2) synthase activity with an IC(50) value of 4.3 +/- 0.2 microM. Furthermore, NP-184 produced a right shift of the concentration-response curve of U46619, indicating a competitive antagonism on TXA(2)/prostaglandin H(2) receptor. Intriguingly, NP-184 also caused a concentration-dependent prolongation of the activated partial thromboplastin time (aPTT) with no changes in the prothrombin and thrombin time, indicating that it selectively impairs the intrinsic coagulation pathway. Oral administration of NP-184 significantly inhibited thrombus formation of the irradiated mesenteric venules in fluorescein sodium-treated mice without affecting the bleeding time induced by tail transection. However, after oral administration, NP-184 inhibited the ex vivo mouse platelet aggregation triggered by collagen and U46619 and also prolonged aPTT. Taken together, the dual antiplatelet and anticoagulant activities of NP-184 may have therapeutic potential as an oral antithrombotic agent in the treatment of thromboembolic disorders.
Cloning and Expression of Pathogenesis-related Protein 4 from Jelly Fig (Ficus Awkeotsang Makino) Achenes Associated with Ribonuclease, Chitinase and Anti-fungal Activities Plant Physiology and Biochemistry : PPB / SociÃ©tÃ© FranÃ§aise De Physiologie VÃ©gÃ©tale. Jul, 2012 | Pubmed ID: 22579939 A cDNA fragment (FaPR4) encoding a class I pathogenesis-related protein 4 (PR-4) from Ficus awkeotsang was obtained by PCR cloning. Plant PR-4s were grouped into class I and II, differing by the presence of ChtBD and hinge. The predicted mature FaPR4 comprises N-terminal chitin-binding domain (ChtBD), hinge, Barwin domain and C-terminal extension. FaPR4-C, an N-terminal truncated form of FaPR4, was designed to mimic the structural feature of class II PR-4s. FaPR4 and FaPR4-C were over-expressed in yeast Pichia pastoris, and both recombinants exhibited RNase and anti-fungal activities. To our knowledge, it is the first report that FaPR4, a member of class I PR-4s has RNase activity as class II. FaPR4 possesses better anti-fungal activities toward Fusarium oxysporum and Sclerotium rolfsii than FaPR4-C. Heat-treated FaPR4 remained RNase and anti-fungal activities; while heat-treated FaPR4-C lost those activities. Therefore, ChtBD of FaPR4 may not only contribute to its anti-fungal but also improve the thermal stability of protein. It also implied the correlation of RNase activity with anti-fungal activity of FaPR4-C. Furthermore, FaPR4 was detected to have weak but significant chitinase activity, and its chitinase activity was reduced after heat treatment. The chitinase activity by FaPR4-C was much lower than FaPR4.
Gene Therapy Rescues Cilia Defects and Restores Olfactory Function in a Mammalian Ciliopathy Model Nature Medicine. Sep, 2012 | Pubmed ID: 22941275 Cilia are evolutionarily conserved microtubule-based organelles that are crucial for diverse biological functions, including motility, cell signaling and sensory perception. In humans, alterations in the formation and function of cilia manifest clinically as ciliopathies, a growing class of pleiotropic genetic disorders. Despite the substantial progress that has been made in identifying genes that cause ciliopathies, therapies for these disorders are not yet available to patients. Although mice with a hypomorphic mutation in the intraflagellar transport protein IFT88 (Ift88Tg737Rpw mice, also known as ORPK mice)5 have been well studied, the relevance of IFT88 mutations to human pathology is unknown. We show that a mutation in IFT88 causes a hitherto unknown human ciliopathy. In vivo complementation assays in zebrafish and mIMCD3 cells show the pathogenicity of this newly discovered allele. We further show that ORPK mice are functionally anosmic as a result of the loss of cilia on their olfactory sensory neurons (OSNs). Notably, adenoviral-mediated expression of IFT88 in mature, fully differentiated OSNs of ORPK mice is sufficient to restore ciliary structures and rescue olfactory function. These studies are the first to use in vivo therapeutic treatment to reestablish cilia in a mammalian ciliopathy. More broadly, our studies indicate that gene therapy is a viable option for cellular and functional rescue of the complex ciliary organelle in established differentiated cells.